Functional Analysis of the N-Acetylglucosamine Metabolic Genes of Streptomyces coelicolor and Role in Control of Development and Antibiotic Production

in Journal of Bacteriology (2012), 194(5), 1136-44

Extracellular Sugar-Phosphates are Assimilated by Streptomyces in a PhoP-dependent Manner

in Antonie van Leeuwenhoek (2012)

The response to copper stress in Streptomyces lividans extends beyond genes under the direct control of a Copper sensitive operon Repressor protein (CsoR)

in Journal of Biological Chemistry (2012)

A Copper sensitive operon Repressor protein has been identified in Streptomyces livdians (CsoRSl) and found to regulate copper homeostasis with attomolar affinity for Cu(I). Solution studies reveal apo ... [more ▼]

A Copper sensitive operon Repressor protein has been identified in Streptomyces livdians (CsoRSl) and found to regulate copper homeostasis with attomolar affinity for Cu(I). Solution studies reveal apo- and CuI-CsoRSl to be a tetramer assembly and a 1.7 Å resolution crystal structure of apo-CsoRSl reveals that a significant conformational change is necessary to enable Cu(I) binding. In silico prediction of the CsoR regulon was confirmed in vitro (EMSA) and in vivo (RNA-seq) which highlighted that next to the csoR gene itself, the regulon consists of two Cu(I) efflux systems involving a CopZ-like copper metallochaperone protein and a CopA P1-type ATPase. While deletion of csoR has only minor effects on S. lividans development when grown under high copper concentrations, mutations of the Cu(I) ligands decrease tolerance to copper as a result of the Cu(I)-CsoR mutants failing to disengage from the DNA targets, thus inhibiting the derepression of the regulon. RNA-seq experiments carried out on samples incubated with exogenous copper and a ΔcsoR strain showed that the set of genes responding to copper stress is much wider than anticipated and largely extends beyond genes targeted by CsoR. This suggests more control levels are operating and directing other regulons in copper homeostasis beside the CsoR regulon [less ▲]

Role of the phenazine-inducing protein Pip in stress resistance of Pseudomonas chlororaphis

in Microbiology (2011), 157(2), 398-407

Chapter 1 Variation in Form and Function The Helix-Turn-Helix Regulators of the GntR Superfamily

in Advances in Applied Microbiology (2009), Chapitre 1

One of the most abundant and widely distributed groups of Helix-turn-helix (HTH) transcription factors is the metabolite-responsive GntR family of regulators (>8500 members in the Pfam database; Jan 2009 ... [more ▼]

One of the most abundant and widely distributed groups of Helix-turn-helix (HTH) transcription factors is the metabolite-responsive GntR family of regulators (>8500 members in the Pfam database; Jan 2009). These proteins contain a DNA-binding HTH domain at the N terminus of the protein and an effector-binding and/or oligomerisation domain at the C terminus, where upon on binding an effector molecule, a conformational change occurs in the protein which influences the DNA-binding properties of the regulator resulting in repression or activation of transcription. This review summarises what we know about the distribution, structure, function and classification of these regulators and suggests that they may have a future role in biotechnology. [less ▲]

Feast or famine: the global regulator DasR links nutrient stress to antibiotic production by Streptomyces

in EMBO Reports (2008), 9(7), 670-675

Members of the soil-dwelling prokaryotic genus Streptomyces produce a wealth of secondary metabolites, including antibiotics and anti-tumour agents. Their formation is intimately coupled with the onset of ... [more ▼]

Members of the soil-dwelling prokaryotic genus Streptomyces produce a wealth of secondary metabolites, including antibiotics and anti-tumour agents. Their formation is intimately coupled with the onset of development, triggered by the nutrient status of the habitat. We propose the first complete signalling cascade from nutrient sensing to development and antibiotic biosynthesis. We show that a high concentration of N-acetylglucosamine - perhaps mimicking the accumulation of N-acetylglucosamine after autolytic degradation of the vegetative mycelium - is a major checkpoint for the onset of secondary metabolism. The response is transmitted to the antibiotic pathway-specific activators via the pleiotropic transcriptional repressor DasR, whose regulon also includes all N-acetylglucosamine-related catabolic genes. The results have led to a new strategy for activating “cryptic” pathways for secondary metabolite biosynthesis, which are abundant in actinomycete genomes, thereby offering new prospects in the fight against multiply drug resistant pathogens and cancers. [less ▲]

PREDetector: A new tool to identify regulatory elements in bacterial genomes

in Biochemical and Biophysical Research Communications (2007), 357(4), 861-864

In the post-genomic area, the prediction of transcription factor regulons by position weight matrix-based programmes is a powerful approach to decipher biological pathways and to modelize regulatory ... [more ▼]

In the post-genomic area, the prediction of transcription factor regulons by position weight matrix-based programmes is a powerful approach to decipher biological pathways and to modelize regulatory networks in bacteria. The main difficulty once a regulon prediction is available is to estimate its reliability prior to start expensive experimental validations and therefore trying to find a way how to identify true positive hits from an endless list of potential target genes of a regulatory protein. Here we introduce PREDetector (Prokaryotic Regulatory Elements Detector), a tool developed for predicting regulons of DNA-binding proteins in bacterial genomes that, beside the automatic prediction, scoring and positioning of potential binding sites and their respective target genes in annotated bacterial genomes, it also provides an easy way to estimate the thresholds where to find reliable possible new target genes. PREDetector can be downloaded freely at http://www.montefiore.ulg.ac.be/-hiard/PreDetector (c) 2007 Published by Elsevier Inc. [less ▲]

Conserved cis-acting elements upstream of genes composing the chitinolytic system of streptomycetes are DasR-responsive elements

in Journal of Molecular Microbiology and Biotechnology (2007), 12(1-2), 60-66

For soil-dwelling bacteria that usually live in a carbon-rich and nitrogen-poor environment, the ability to utilize chitin -the second most abundant polysaccharide on earth is a decisive evolving ... [more ▼]

For soil-dwelling bacteria that usually live in a carbon-rich and nitrogen-poor environment, the ability to utilize chitin -the second most abundant polysaccharide on earth is a decisive evolving advantage as it is a source for both elements. Streptomycetes are high-GC Gram-positive soil bacteria that are equipped with a broad arsenal of chitinase-degrading genes. These genes are induced when the streptomycetes sense the presence of chitooligosaccharides. Their expression is repressed as soon as more readily assimilated carbon sources become available. This includes for example glucose or N-acetylglucosamine, the monomer subunit of chitin. Historically, the first cis-acting elements involved in carbon regulation in streptomycetes were found more than a decade ago upstream of chitinase genes, but the transcriptional regulator had so far remained undiscovered. In this work, we show that these cis-acting elements consist of inverted repeats with multiple occurrences and are bound by the HutC/GntR type regulator DasR. We have therefore designated these sites as DasR-responsive elements (dre). DasR, which is also the repressor of the genes for the N-acetylglucosamine-specific phosphotransferase transport system, should therefore play a critical role in sensing the balance between the monomeric and polymeric forms of N-acetylglucosamine. Copyright (c) 2007 S. Karger AG, Basel. [less ▲]

DevA, a GntR-like transcriptional regulator required for development in Streptomyces coelicolor

in Journal of Bacteriology (2006), 188(14), 5014-5023

The gram-positive filamentous bacterium Streptomyces coelicolor has a complex developmental cycle with three distinct phases: growth of the substrate mycelium, development of reproductive structures ... [more ▼]

The gram-positive filamentous bacterium Streptomyces coelicolor has a complex developmental cycle with three distinct phases: growth of the substrate mycelium, development of reproductive structures called aerial hyphae, and differentiation of these aerial filaments into long chains of exospores. During a transposon mutagenesis screen, we identified a novel gene (devA) required for proper development. The devA mutant produced only rare aerial hyphae, and those that were produced developed aberrant spore chains that were much shorter than wild-type chains and had misplaced septa. devA encodes a member of the GntR superfamily, a class of transcriptional regulators that typically respond to metabolite effector molecules. devA forms an operon with the downstream gene devB, which encodes a putative hydrolase that is also required for aerial mycelium formation on R5 medium. S1 nuclease protection analysis showed that transcription from the single devA promoter was temporally associated with vegetative growth, and enhanced green fluorescent protein transcriptional fusions showed that transcription was spatially confined to the substrate hyphae in the wild type. In contrast, devAB transcript levels were dramatically upregulated in a devA mutant and the devA promoter was also active in aerial hyphae and spores in this background, suggesting that DevA might negatively regulate its own production. This suggestion was confirmed by gel mobility shift assays that showed that DevA binds its own promoter region in vitro. [less ▲]

Pip, a novel activator of phenazine biosynthesis in Pseudomonas chlororaphis PCL1391

in Journal of Bacteriology (2006), 188(23), 8283-8293

Secondary metabolites are important factors for interactions between bacteria and other organisms. Pseudomonas chlororaphis PCL1391 produces the antifungal secondary metabolite phenazine-1-carboxamide ... [more ▼]

Secondary metabolites are important factors for interactions between bacteria and other organisms. Pseudomonas chlororaphis PCL1391 produces the antifungal secondary metabolite phenazine-1-carboxamide (PCN) that inhibits growth of Fusarium oxysporum f. sp. radius lycopersici the causative agent of tomato foot and root rot. Our previous work unraveled a cascade of genes regulating the PCN biosynthesis operon, phzABCDEFGH. Via a genetic screen, we identify in this study a novel TetR/AcrR regulator, named Pip (phenazine inducing protein), which is essential for PCN biosynthesis. A combination of a phenotypical characterization of a pip mutant, in trans complementation assays of various mutant strains, and electrophoretic mobility shift assays identified Pip as the fifth DNA-binding protein so far involved in regulation of PCN biosynthesis. In this regulatory pathway, Pip is positioned downstream of PsrA (Pseudomonas sigma factor regulator) and the stationary-phase sigma factor RpoS, while it is upstream of the quorum-sensing system PhzI/PhzR. These findings provide further evidence that the path leading to the expression of secondary metabolism gene clusters in Pseudomonas species is highly complex. [less ▲]